PROJECT SUMMARY/ABSTRACT Ischemia during myocardial infarction (MI) causes sudden cardiomyocyte death that initiates an intense inflammatory response, but timely resolution of the inflammatory phase is necessary to move into the reparative phase and minimize scarring. A prolonged pro-inflammatory phase is associated with an adverse ventricular remodeling response that can progress to clinical heart failure. Current therapeutic options are limited and fail to prevent or reverse immune-driven fibrosis after myocardial infarction. The theme of the proposed work is that IL- 33 driven modulation of the immune response can minimize fibrosis and preserve myocardial function after ischemic injury. Bioscaffolds composed of extracellular matrix (ECM) have been shown to minimize scarring after MI in rodent models, but the underlying mechanisms are only partially understood. The identification of interleukin-33 (IL-33) stably stored within the ECM suggests a potential role in the ECM-mediated immunomodulation and constructive remodeling response. Emerging evidence shows that IL-33 supports tissue repair, especially in models of cardiovascular disease, by signaling through its cognate receptor, ST2. IL-33 has previously been considered a nuclear protein, but recent studies have found IL-33 stably integrated into the ECM by encapsulation within matrix bound nanovesicles (MBV) that are released during ECM degradation to target infiltrating immune cells. We have found that MBV containing IL-33 can direct macrophage differentiation toward a pro-remodeling M2 phenotype through a pathway that does not involve the ST2 receptor. The identification of an ST2-independent IL-33 signaling mechanism induced by MBV is novel in itself but is particularly important in the context of myocardial infarction because expression levels of a soluble, antagonistic form of ST2 are increased following MI injury and associated with poor prognosis. It is hypothesized that MBV within the ECM are a critical source of IL-33 necessary to mediate early ST2-independent pro-remodeling macrophage differentiation and direct constructive remodeling after myocardial ischemia. The proposed studies will first define the repertoire of genes regulated by the ST2-independent signaling mechanism in macrophages and then investigate the role of IL-33 mediated immunomodulation in preventing immune-driven fibrosis. The results of this work my enable novel immunomodulatory approaches to protect the myocardium after ischemic injury.